Project Summary/Abstract: Proteome-wide protein methylation is an epigenetic event that is regulated by >60 protein methyltransferases (PMTs). Because conventional approaches can only uncover a small fraction of PMT substrates without knowing PMT methylome under defined cellular settings, we have only partial insight into epigenetic role(s) of PMT in many essential biological settings. Selective perturbation of PMTs is also of great use to elucidate their role(s). While PMT- associated biology is often probed via genetic perturbation, this approach, unlike small-molecule inhibitors, lacks temporal (turn on/off), spatial (loci-specific) and dose (a range of efficacy) controls. In addition, the outcomes of genetic versus pharmacological perturbation of PMTs can be different because the former completely ablates a PMT protein whereas the latter only affects a subset of PMT's functions. However, developing selective and potent small-molecule inhibitors of PMTs is challenging. The objective of this proposal is to integrate novel chemical tools (high-quality inhibitors and bioorthogonal activity probes of PMTs) with conventional methods to examine the outcome(s) and therapeutic potential of pharmacological inhibition of a PMT. We plan to define the methylation events critical for genome replication and DNA damage responses, pharmacologically manipulate them in a temporal and dose-dependent manner, and predict its outcomes using accurate animal models.